Accumulating evidence has demonstrated that FHIT (fragile histidine triad) is a bona fide tumour suppressor gene in a large fraction of human tumours, including hepatocellular cancer. A virus-based delivery system has been developed to transfer the FHIT gene into many types of cancer cells to inhibit growth or even induce apoptosis. However, a protein-based replacement strategy for FHIT has not been performed in cancer cells. Here, we used HIV-TAT (transactivator of transcription)-derived peptide to transfer the purified FHIT protein into HCC (hepatocellular carcinoma) cells and determine the biological effect of this fusion protein in inducing apoptosis. Affinity chromatography was used to purify TAT peptide-fused human FHIT (TAT–FHIT) protein from BL21 Escherichia coli. Immunofluorescence staining and Western blot analysis were performed to identify the expression and internalization of TAT–FHIT in HCC cells compared with the purified FHIT protein. Our study showed that TAT–FHIT protein can translocate into cancer cells in 1 h after incubation at 37°C. Furthermore, the results of MTT [3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-2H-tetrazolium bromide] assay, Annexin-V staining and Western blotting demonstrated that TAT–FHIT can robustly inhibit growth and induce apoptosis of HCC cells in vitro. In addition, a mechanistic study showed that both exogenous and intrinsic apoptotic pathways were involved in TAT–FHIT-mediated apoptosis and this effect could be attenuated partially by a mitochondrial protector TAT-BH4, indicating that mitochondrion plays a critical role in TAT–FHIT-mediated pro-apoptotic effect in cancer cells. Taken together, our study suggests that TAT–FHIT is a potential pro-apoptotic molecule in HCC cells and strengthen the hypothesis of its therapeutic application against HCC.
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Research Article|
January 30 2012
HIV-TAT-fused FHIT protein functions as a potential pro-apoptotic molecule in hepatocellular carcinoma cells
Gui-Rong Yu;
Gui-Rong Yu
1
*State Key Laboratory of Cancer Biology, Department of Biochemistry and Molecular Biology, Fourth Military Medical University, 710032 Xi'an, People's Republic of China
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Wei-Wei Qin;
Wei-Wei Qin
1
*State Key Laboratory of Cancer Biology, Department of Biochemistry and Molecular Biology, Fourth Military Medical University, 710032 Xi'an, People's Republic of China
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Ji-Peng Li;
Ji-Peng Li
1
*State Key Laboratory of Cancer Biology, Department of Biochemistry and Molecular Biology, Fourth Military Medical University, 710032 Xi'an, People's Republic of China
†Department of Gastrointestinal Surgery, Xijing Hospital, Fourth Military Medical University, 710032 Xi'an, People's Republic of China
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Wei Hua;
Wei Hua
1
‡Department of Obstetrics and Gynecology, Xijing Hospital, Fourth Military Medical University, 710032 Xi'an, People's Republic of China
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Yan-Ling Meng;
Yan-Ling Meng
§Department of Immunology, Fourth Military Medical University, 710032 Xi'an, People's Republic of China.
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Rui Chen;
Rui Chen
*State Key Laboratory of Cancer Biology, Department of Biochemistry and Molecular Biology, Fourth Military Medical University, 710032 Xi'an, People's Republic of China
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Bo Yan;
Bo Yan
*State Key Laboratory of Cancer Biology, Department of Biochemistry and Molecular Biology, Fourth Military Medical University, 710032 Xi'an, People's Republic of China
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Lei Wang;
Lei Wang
*State Key Laboratory of Cancer Biology, Department of Biochemistry and Molecular Biology, Fourth Military Medical University, 710032 Xi'an, People's Republic of China
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Xiang Zhang;
Xiang Zhang
*State Key Laboratory of Cancer Biology, Department of Biochemistry and Molecular Biology, Fourth Military Medical University, 710032 Xi'an, People's Republic of China
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Lin-Tao Jia;
Lin-Tao Jia
*State Key Laboratory of Cancer Biology, Department of Biochemistry and Molecular Biology, Fourth Military Medical University, 710032 Xi'an, People's Republic of China
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Jing Zhao;
Jing Zhao
*State Key Laboratory of Cancer Biology, Department of Biochemistry and Molecular Biology, Fourth Military Medical University, 710032 Xi'an, People's Republic of China
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Rui Zhang;
Rui Zhang
2
*State Key Laboratory of Cancer Biology, Department of Biochemistry and Molecular Biology, Fourth Military Medical University, 710032 Xi'an, People's Republic of China
2Correspondence may be addressed to either of these authors (email agyang@fmmu.edu.cn or ruizhang@fmmu.com.cn).
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An-Gang Yang
An-Gang Yang
2
*State Key Laboratory of Cancer Biology, Department of Biochemistry and Molecular Biology, Fourth Military Medical University, 710032 Xi'an, People's Republic of China
§Department of Immunology, Fourth Military Medical University, 710032 Xi'an, People's Republic of China.
2Correspondence may be addressed to either of these authors (email agyang@fmmu.edu.cn or ruizhang@fmmu.com.cn).
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Publisher: Portland Press Ltd
Received:
March 15 2011
Revision Received:
June 10 2011
Accepted:
June 16 2011
Accepted Manuscript online:
June 16 2011
Online ISSN: 1573-4935
Print ISSN: 0144-8463
© The Authors Journal compilation © 2012 Biochemical Society
2012
Biosci Rep (2012) 32 (3): 271–279.
Article history
Received:
March 15 2011
Revision Received:
June 10 2011
Accepted:
June 16 2011
Accepted Manuscript online:
June 16 2011
Citation
Gui-Rong Yu, Wei-Wei Qin, Ji-Peng Li, Wei Hua, Yan-Ling Meng, Rui Chen, Bo Yan, Lei Wang, Xiang Zhang, Lin-Tao Jia, Jing Zhao, Rui Zhang, An-Gang Yang; HIV-TAT-fused FHIT protein functions as a potential pro-apoptotic molecule in hepatocellular carcinoma cells. Biosci Rep 1 June 2012; 32 (3): 271–279. doi: https://doi.org/10.1042/BSR20110033
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